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PLANT TISSUE CULTURE PRESENTATION

md.Nur uddin titu


Bsc In biotechnology &
genetic Engineering
NSTU.
Reference:
Gamborg O.L., Miler R.A. And Ojima K.,
1968. page no:151-158.

Plant tissue culture


Plant tissue culture
• Plant tissue culture is a collection of techniques
used to maintain or grow plant cells, tissues or
organs under sterile conditions on a
nutrient culture medium of known composition.
• Plant tissue culture is widely used to produce
clones of a plant in a method known as
micropropagation
BASIC REQUIREMENT OF A PLANT
TISSUE CULTURE MEDIA
1. Complex mixture of salts: Essential elements, or
mineral ions.
2. Organic supplement: vitamins and/or amino acids.
3. Carbon source: usually sugar sucrose.
4. Gelling agents
5. Plant Growth Regulators
6. Antibiotics
1. Complex mixture of salts: Essential
elements, or mineral ions.
The essential elements can further be divided
into the following categories:

a. Macroelements (or macronutrients)

b. Microelements (or micronutrients)

c. Iron Source
Macroelements:
• These elements are required in large amounts
for plant growth and development.
Ex :
Nitrogen, phosphorus, potassium,
magnesium, calcium etc..
Microelements:
These elements are required in trace amounts for
plant growth and development.
Ex :
Manganese, iodine, copper, cobalt, boron,
molybdenum, iron
Organic supplements:
• These include vitamins and amino acids. Two
vitamins, i.e., thiamine (vitamin B1) and
myoinositol (a vitamin B) are essential for the
culture of plant cells in vitro.

• The most commonly used amino acid is glycine.


Carbon source:
• The most commonly used carbon source is
sucrose.
Gelling agents:
• Plant tissue culture media can be used in either
liquid or 'solid' forms, depending on the type of
culture being grown.
• Agar, produced from seaweed, is the most common
type of gelling agent .
Plant growth regulators:
• Specific media manipulations can be used to direct
the development of plant cells in culture due to
plasticity and totipotency.
Plant growth regulators:
• There are five main classes of plant growth regulator
used in plant cell culture, namely:
a. Auxins
b. Cytokinins
c. Gibberellins
d. Abscisic acid
e. Ethylene
WHITES AND GAMBORG’S
MEDIA
Use
Medium is used for callus culture and cell
suspension culture
Summary
• established by Gamborg O.L. (1968) for callus and
cellsuspension culture of Glycine max (Family-
Fabaceae).
• This medium is widely used for in vitro plant cell,
tissue and organ culture.
• Principle

• Gamborg Medium provides all essential


Macroelements, Microelements, &Vitamins for the
growth of plant cell, tissue and organ culture
invitro.

• The medium generally contains greater proportion


of nitrate and potassium but low concentration of
ammonia
Media composition (gamborgs)
Macroelements mg/L

• Potassium nitrate 2500.00


• Ammonium sulphate 134.00
• Calcium chloride anhydrous 113.24
• Magnesium sulphate 122..09
• Sodium phosphate monobasic 130.42
These elements are required in large amounts
for plant growth and development.
• Need of macro nutrients is higher.

• It is present in milli molar (mM) quantities


(more than 30 ppm/1 or mg/1)

• Macro nutrients provide both anions and


cations for the plant cells
Phosphorus
• Very important for energy metabolism.
• Essential element for DNA & RNA.
• Deficiency may cause delayed growth and dark green
coloure of leaves.
• Supplied instead of sodium hydrogen phosphate or
potassium hydrogen phosphate.
Magnesium
• Essential for enzymatic reactions, energy metabolism(ATP synthesis).

• Supplied as magnesium sulphate.


sulphur
• Important substance.
• It increases the cholophyll in leaves.
• Supplied as magnesium Sulphate and
Potassium Sulphate.
Microelements mg/L
• Manganese sulphate. 10.00
• Boric Acid 3.00
• Potassium iodide 0.75
• Molybdic acid (sodium salt). 0.21
• Zinc sulphate. 2.00
• Copper sulphate. 0.025
• Cobalt chloride. 0.025
• Ferrous sulphate. 27.80
• Na .EDTA 37.26
These elements are required in trace amounts
for plant growth and development.
Zink
• Zn plays an active role in protein synthesis .
• Supplied as Zinc Sulphate

Manganese
• Plays an important role in the Hill reaction of
photasynthesis.
• Required in many enzymatic activities.
• Supplied as Manganese Sulphate.
Copper
• Copper plays important role in photosynthesis.
• Supplied as Copper Sulphate.

Hexitols

• Most tissue culture media have this compound.

• Essential for seed germination, sugar transport,


carbohydrate metabolism, membrane structure
and cell wall formation
Vitamins mg/L
• Myo-Inositol 100.00
• Thiamine HCL 10.00
• Pyridoxine HCL 1.00
• Nicotinic acid (Free acid) 1.00
Thiamine content had been increased in the
medium which supported the growth of cell
suspension culture.
The medium lacks glycine.
Gelling Agent
• Agar 8000.00
TOTAL: 23.23 gm/litre
Whites media

• White medium was established by white (1963)


for root culture of tomato
• This is one of the earliest media with low salt
formulation
• Nitrate concentration is 19 % less than MS media
• Prominence was given to MgSO4
• White Medium Composition :
• Ingredients milligrams/litre
• Potassium nitrate 80.00
• Calcium nitrate 221.96
• Magnesium sulphate 360.00
• Sodium phosphate monobasic 18.98
• Potassium chloride 65.00
• Sodium sulphate 200.00
• Manganese sulphate.H2O 5.04
• Boric acid 1.50
• Potassium iodide 0.75
• Molybdenum trioxide 0.001
• Zinc sulphate.7H2O 2.67
• Copper sulphate.5H2O 0.01
• Ferrous sulphate.7H2O 2.50
• myo - Inositol 100.00
• Thiamine hydrochloride 0.10
• Pyridoxine hydrochloride 0.10
• Nicotinic acid (Free acid) 0.50
• Glycine (Free base) 3.00
• Sucrose 20000.00
• TOTAL gm/litre 21.06
THANK U….

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